Stimulatory effect of NH4+ on the transport of leucine and glucose in an anaerobic alkaliphile

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Abstract
An anaerobic alkaliphile, EP01, specifically requires NH4+ for the acceleration of amino acid and glucose transport [Koyama, N. (1988) FEBS Lett. 253, 187-189]. In this paper, we attempted to clarify how NH4+ is involved in the transport system. The bacterium acidifies the cytoplasm, which was suggested to result in NH4+ accumulation when NH4Cl was added to the medium. Increase of the NH4Cl concentration administered to the medium caused the acceleration of leucine and glucose transport, which was accompanied by an increase in the internal pH and the absolute internal concentration of NH4+, whereas a decrease in the concentration ratio of internal NH4+/external NH4+ was observed. The addition of 3 mM NH4Cl, which resulted in significant stimulation of leucine and glucose transport, raised the internal NH4+ concentration by 42 mM, but the internal pH only by 0.1 units. It seems more likely that leucine and glucose transport are accelerated depending on the increase in the internal NH4+ concentration rather than the increase in the internal pH. By the imposition of an inwardly directed Na+ gradient, the K(+)-loaded membrane vesicles accumulated leucine and glucose, indicating that a sodium chemical potential is available for active transport. The membrane of the bacterium exhibited a Na(+)-stimulated ATPase activity which was remarkably enhanced by the addition of NH4Cl, depending on its concentration, and was inhibited by vanadate. Leucine and glucose transport were inhibited by vanadate. Based on these results, we propose a mechanism in which NH4+ contributes internally to leucine and glucose transport, depending on its concentration, by the activation of a Na(+)-translocating ATPase responsible for the generation of a sodium chemical potential.